Understanding molecular mechanisms of somatosensory signaling pathways is critical to find new methods of alleviating pain. To discover novel nociception pathways, we have taken a forward genetic approach using the fruitfly Drosophila melanogaster. Using a larval behavioral response to noxious stimuli we have identified mutants in the painless gene that are defective in mechanical and thermal nociception. Painless is expressed in multidendritic neurons which morphologically resemble vertebrate nociceptive neurons having multiply branched naked neurites beneath the epidermis. The Painless protein is a member of the Transient Receptor Potential family of ion channels, many of which have been implicated in temperature transduction and in mechanotransduction. In Drosophila, three TRPA channels have been implicated as thermosensors;the Painless channel, which we identified genetically as required for thermal nociception, the Drosophila TRPA1 (dTRPAl) channel which has been implicated in thermotaxis, and a third channel named Pyrexia that provides resistance to thermal stress. While it is clear that TRP channels can function in thermosensory and mechanical signal transduction, the molecular mechanisms responsible for mediating responses to temperature and mechanical stimuli by these channels are still a mystery. We are ideally positioned to use in vivo genetics to understand the basic principles underlying TRP channel function in mechanotransduction and in thermotransduction. We will test three hypotheses: 1) that Drosophila TRPA channels combinatorially determine the thermal nociception threshold of the Drosophila larva;2) that a specific subclass of Drosophila multidendritic sensory neuron functions in nociception;and 3) that ankyrin repeat domains of the Painless channel are essential for mechanotransduction. These studies will provide insight into molecular and cellular mechanisms of nociception that will assist in developing therapeutic interventions for the treatment of pain.